Distinct roles for H4 and H2A.Z acetylation in RNA transcription in African trypanosomes

Abstract

Despite histone H2A variants and acetylation of histones occurring in almost every eukaryotic organism, it has been difficult to establish direct functional links between canonical histones or H2A variant acetylation, deposition of H2A variants and transcription. To disentangle these complex interdependent processes, we devised a highly sensitive strategy for quantifying histone acetylation levels at specific genomic loci. Taking advantage of the unusual genome organization in Trypanosoma brucei, we identified 58 histone modifications enriched at transcription start sites (TSSs). Furthermore, we found TSS-associated H4 and H2A.Z acetylation to be mediated by two different histone acetyltransferases, HAT2 and HAT1, respectively. Whereas depletion of HAT2 decreases H2A.Z deposition and shifts the site of transcription initiation, depletion of HAT1 does not affect H2A.Z deposition but reduces total mRNA levels by 50%. Thus, specifically reducing H4 or H2A.Z acetylation levels enabled us to reveal distinct roles for these modifications in H2A.Z deposition and RNA transcription.

Histone Modifications and Other Facets of Epigenetic Regulation in Trypanosomatids: Leaving Their Mark

“…The primary role of H4K10 acetylation in transcription thus appears to be the facilitation of H2A.Z deposition at TSSs. In the absence of H2A.Z, the transcriptional machinery may find an alternate start site in a more conducive location on the template (18). In synchrony with these findings, LdHAT2 depletion also did not have a global impact on transcript levels.…”

“…These included H2A.Z and H2B.V hyperacetylation and H3 methylations. The well characterized H3K76 methylation marks and the H4K4Ac modification were equally detected at TSS and non-TSS nucleosomes, ruling out a role for these two modifications in PolII-mediated transcription (18). Nevertheless, the DOT1B protein that mediates H3K76 trimethylation tightly regulates the silencing and transcriptional switching of PolI-transcribed VSGs, with deletion of DOT1B resulting in the occasional simultaneous expression of two VSGs (73).…”

“…TbHAT1 may also bind to H3K4me3 at the TSSs via its Tudor domain and thereby be anchored for carrying out acetylation of H2A.Z. In the light of data from various groups (18,63,99,101,103,104), it is tempting to speculate on a possible sequence of events at the dSSRs involving cross talk between the different histone PTMs and culminating in transcriptional activation (Fig. 2), though further detailed investigations are needed to support the proposed hypothesis.…”

“…A recent study (published while this article was under review) has identified the histone PTMs associated with the distinctive T. cruzi parasite life forms, with over 60 PTM fluctuations during the life cycle ( 17 ). Two independent studies (one of which was published while this article was under review) have recently unearthed more than 150 PTMs on T. brucei histones also ( 18 , 19 ). Strikingly, virtually no acetylation marks were detectable in H3.V and H4.V.…”

“…Strikingly, virtually no acetylation marks were detectable in H3.V and H4.V. Kraus et al ( 18 ) compared the histone acetylation and methylation marks on nucleosomes at transcription start sites (TSSs) with those on nucleosomes at other sites in T. brucei and found some modifications to be specific to TSSs while others were universally distributed. Histone modifications have been globally identified in Plasmodium and Toxoplasma as well ( 20 , 21 ).…”